Activation of a latent nuclear localization signal in the NH₂ terminus of γ-ENaC initiates feedback regulation of channel activity

Proteolytic enzymes cleave the epithelial Na⁺ channel (ENaC) at several positions releasing, in part, the NH₂ terminus of the γ-subunit. Cleavage increases ENaC activity by increasing open probability; however, the role of polypeptides cleaved from the channel core remains unclear. We find that the cytosolic NH₂ terminus of γ-ENaC unexpectedly targets to the nucleus being particularly strong in nucleoli. In contrast, the cytosolic COOH terminus targets to the cytoplasm and plasma membrane in a manner similar to full-length subunits. Targeting of the cytosolic NH2 terminus of γ-ENaC to the nucleus has functional consequences for coexpression of eGFP-fusion proteins containing this segment of the channel, but not the COOH terminus, decrease ENaC activity in a dose-dependent manner. The mechanism of this negative regulation is associated with a decrease in the functional half-life of ENaC at the plasma membrane. Inspection of the primary amino acid sequence of γ-ENaC reveals possible nuclear localization signals (NLS) conserved at the extreme NH₂ terminus and just preceding the first transmembrane domain. Disruption of the putative NLS preceding the first transmembrane domain in γ-ENaC but not that at the extreme NH₂ terminus abolishes both targeting to the nucleus and negative regulation of ENaC activity. These findings are consistent with the release of the NH2 terminus of γ-ENaC following cleavage being functionally important for signaling to the nucleus in a manner similar to Notch signaling and release of the cytosolic COOH-terminal tail of polycystin-1.